Mathematical model for early development of the sea urchin embryo

In Xenopus and Drosophila, the nucleocytoplasmic ratio controls many aspects of cell-cycle remodeling during the transitory period that leads from fast and synchronous cell divisions of early development to the slow, carefully regulated growth and divisions of somatic cells. After the fifth cleavage in sea urchin embryos, there are four populations of differently sized blastomeres, whose interdivision times are inversely related to size. The inverse relation suggests nucleocytoplasmic control of cell division during sea urchin development as well. To investigate this possibility, we developed a mathematical model based on molecular interactions underlying early embryonic cell-cycle control. Introducing the nucleocytoplasmic ratio explicitly into the molecular mechanism, we are able to reproduce many physiological features of sea urchin development.     

Cell movements in the sea urchin embryo.

Recent studies show that gastrulation in the sea urchin embryo involves movement of cells over the blastopore lip (involution). Some cells in the vegetal plate of the late blastula become bottle-shaped but they play a limited role in gastrulation. The functions of specific integrins, regulators of cell-cell adhesion, and extracellular matrix components in gastrulation are currently being analyzed. In addition, light-microscopic studies continue to provide a unique picture of dynamic cell behavior in vivo.     

Shaping BMP morphogen gradients in the Drosophila embryo and pupal wing

In the early Drosophila embryo, BMP-type ligands act as morphogens to suppress neural induction and to specify the formation of dorsal ectoderm and amnioserosa. Likewise, during pupal wing development, BMPs help to specify vein versus intervein cell fate. Here, we review recent data suggesting that these two processes use a related set of extracellular factors, positive feedback, and BMP heterodimer formation to achieve peak levels of signaling in spatially restricted patterns. Because these signaling pathway components are all conserved, these observations should shed light on how BMP signaling is modulated in vertebrate development.     

Membrane formation and membrane contacts during the development of the sea urchin embryo

Summary Sea urchin embryos, 8-cell stage to pluteus stage, fixed in osmium tetroxide and embedded in Epon 812 were observed by electron microscopy. At no point in the development were syncytial junctions between the embryonic cells found. During the cleavage stages the membrane contact was closer than in later stages. In early blastula stages intercellular clefts appeared which in the gastrula stage demarcate every cell. At the same time a ringshaped desmosome structure develops at the outer cell surface. In the pluteus stage a closer cell contact is re-established. With proceeding embryogenesis endoplasmic membranes will attach to the cell membrane. These membrane structures may even be of nuclear origin. Gradually, long protrusions, vesicles and lamellae begin to be formed from the nuclear membrane. The commencement of this nuclear activity coincides in time with the formation of nucleoli. At cell division the new cell membrane seemed to arise partly independently of the cleavage furrow from a system of cytoplasmic vesicles.The investigation was facilitated by grants from the Nordic Insulin Foundation.I am indebted to Dr. Torsten Olsson and Miss Brita Nilsson for procuring the material and to Mrs. Mariann Carleson for technical aid.     

Determination and morphogenesis in the sea urchin embryo

The study of the sea urchin embryo has contributed importantly to our ideas about embryogenesis. This essay re-examines some issues where the concerns of classical experimental embryology and cell and molecular biology converge. The sea urchin egg has an inherent animal-vegetal polarity. An egg fragment that contains both animal and vegetal material will produce a fairly normal larva. However, it is not clear to what extent the oral-aboral axis is specified in embryos developing from meridional fragments. Newly available markers of the oral-aboral axis allow this issue to be settled. When equatorial halves, in which animal and vegetal hemispheres are separated, are allowed to develop, the animal half forms a ciliated hollow ball. The vegetal half, however, often forms a complete embryo. This result is not in accord with the double gradient model of animal and vegetal characteristics that has been used to interpret almost all defect, isolation and transplantation experiments using sea urchin embryos. The effects of agents used to animalize and vegetalize embryos are also due for re-examination. The classical animalizing agent, Zn2+, causes developmental arrest, not expression of animal characters. On the other hand, Li+, a vegetalizing agent, probably changes the determination of animal cells. The stability of these early determinative steps may be examined in dissociation-reaggregation experiments, but this technique has not been exploited extensively. The morphogenetic movements of primary mesenchyme are complex and involve a number of interactions. It is curious that primary mesenchyme is dispensable in skeleton formation since in embryos devoid of primary mesenchyme, the secondary mesenchyme cells will form skeletal elements. It is likely that during its differentiation the primary mesenchyme provides some of its own extracellular microenvironment in the form of collagen and proteoglycans. The detailed form of spicules made by primary mesenchyme is determined by cooperation between the epithelial body wall, the extracellular material and the inherent properties of primary mesenchyme cells. Gastrulation in sea urchins is a two-step process. The first invagination is a buckling, the mechanism of which is not understood. The secondary phase in which the archenteron elongates across the blastocoel is probably driven primarily by active cell repacking. The extracellular matrix is important for this repacking to occur, but the basis of the cellular-environmental interaction is not understood.(ABSTRACT TRUNCATED AT 400 WORDS)     

Fibronectin in the developing sea urchin embryo

The presence of fibronectin in developing sea urchin embryos was studied uing immunofluorescence staining. The fluorescence pattern indicates that fibronectin is found on the cell surfaces and between cells in the blastula and gastrula stages, indicating that it plays a role in cell adhesion. Its presence on invaginating cells also suggests its involvement in morphogenesis during early development.     

Metabolic interconversion of dolichol and dolichyl phosphate during development of the sea urchin embryo

The in vivo and in vitro synthesis and turnover of dolichol and dolichyl phosphate have been studied over the course of early development in sea urchin embryos. Synthesis of dolichol and dolichyl phosphate was studied in vivo and in vitro using [3H]acetate and [14C] isopentenylpyrophosphate, respectively, as precursors. Both the in vivo and in vitro results indicate that the principal labeled end product of de novo synthesis is the free alcohol, and that this alcohol is subsequently phosphorylated to produce dolichyl phosphate. The presence of 30 microM compactin inhibits the de novo synthesis of dolichol from [3H]acetate by greater than 90%, but has no effect on the incorporation of 32Pi into dolichyl phosphate for more than 6 h, thus suggesting that during this time interval the major source of dolichyl phosphate is preformed dolichol. The rate of turnover of the [3H]acetate-labeled polyisoprenoid backbone of dolichyl phosphate is very slow (t1/2 = 40-70 h). In contrast, the rate of loss of the [32P]phosphate headgroup is more rapid (t1/2 = 5.7-7.7 h) and increases over the course of development. Finally, dolichyl phosphate phosphatase activity has been measured in vitro. The activity of this enzyme, which can be distinguished from phosphatidic acid phosphatase, was found to increase as a function of development, in qualitative agreement with the increased turnover of 32P from dolichyl phosphate observed in vivo. These results suggest that the phosphate moiety of dolichyl phosphate is in a dynamic state, and that dolichol kinase and dolichyl phosphate phosphatase play key roles in regulating the cellular level of dolichyl phosphate.     

Secondary mesenchyme of the sea urchin embryo: ontogeny of blastocoelar cells.

Secondary mesenchyme in sea urchin embryos is released into the blastocoel after primary mesenchyme, and although these cells have been recognized for some time, we lack knowledge about many fundamental aspects of their origin and fate. Here we documented the ontogeny of one of the principal, and least well-known, types of cells derived from secondary mesenchyme. The blastocoelar cells arise from mesenchyme released from the tip of the archenteron following the initial phase of gastrulation. The cells migrate with their cell bodies suspended in the blastocoel, rather than being apposed to the basal lamina like primary mesenchyme. The cells extend numerous fine filopodia to form a network of cytoplasmic processes around the gut, along the skeletal rods, and within the larval arms. Once the network is formed, the cells maintain their positions, although they actively translocate vesicles and cytoplasm along their filopodia. Cell counts indicate there is an initial recruitment of cells during gastrulation, followed by a more gradual increase in cell number after the larva begins to feed. Lineage studies in which 16-cell-stage macromeres were injected with horseradish peroxidase indicate that almost all of the macromere-derived mesenchyme forms pigment cells and blastocoelar cells. We propose that blastocoelar cells are a distinct subset of secondary mesenchyme that forms fibroblast-like cells in the blastocoel of sea urchin embryos.     

Matrix metalloproteases of the developing sea urchin embryo

Abstract. A distinct group of metalloproteases has been identified in the developing sea urchin embryo by gelatin substrate gel zymography, a highly sensitive protease detection assay. The developing Arbacia embryo exhibited four prominent bands of gelatinase activity with apparent molecular masses of 55, 50, 42 and 38 kDa. The activity of the 55, 42 and 38 kDa tissue gelatinases increased and that of the 50 kDa tissue gelatinase decreased during embryonic development. All four enzymes were EDTA- and 1,10-phenanthroline sensitive and phenyl methyl sulphonyl fluoride (PMSF) insensitive. None of the enzymes had detectable caseinolytic activity in casein substrate gels. Although the Arbacia enzymes possessed a number of properties that are characteristic of the mammalian matrix metalloprotease family, they did not appear to be converted to lower molecular weight forms by organomercurial treatment and are distinct in this aspect. The Arbacia metalloproteases are candidate enzymes for the tissue and matrix remodeling that occurs during sea urchin embryo development.     

Matrix metalloproteases of the developing sea urchin embryo

Abstract. A distinct group of metalloproteases has been identified in the developing sea urchin embryo by gelatin substrate gel zymography, a highly sensitive protease detection assay. The developing Arbacia embryo exhibited four prominent bands of gelatinase activity with apparent molecular masses of 55, 50, 42 and 38 kDa. The activity of the 55, 42 and 38 kDa tissue gelatinases increased and that of the 50 kDa tissue gelatinase decreased during embryonic development. All four enzymes were EDTA- and 1,10-phenanthroline sensitive and phenyl methyl sulphonyl fluoride (PMSF) insensitive. None of the enzymes had detectable caseinolytic activity in casein substrate gels. Although the Arbacia enzymes possessed a number of properties that are characteristic of the mammalian matrix metalloprotease family, they did not appear to be converted to lower molecular weight forms by organomercurial treatment and are distinct in this aspect. The Arbacia metalloproteases are candidate enzymes for the tissue and matrix remodeling that occurs during sea urchin embryo development.     

Cell lineage conversion in the sea urchin embryo

The mesoderm of the sea urchin embryo conventionally is divided into two populations of cells; the primary mesenchyme cells (PMCs), which produce the larval skeleton, and the secondary mesenchyme cells (SMCs), which differentiate into a variety of cell types but do not participate in skeletogenesis. In this study we examine the morphogenesis of embryos from which the PMCs have been removed microsurgically. We confirm the observation of Fukushi (1962) that embryos lacking PMCs form a complete skeleton, although in a delayed fashion. We demonstrate by microsurgical and cell marking experiments that the appearance of skeletogenic cells in such PMC-deficient embryos is due exclusively to the conversion of other cells to the PMC phenotype. Time-lapse video recordings of PMC-deficient embryos indicate that the converting cells are a subpopulation of late-ingressing SMCs. The conversion of these cells to the skeletogenic phenotype is accompanied by their de novo expression of cell surface determinants normally unique to PMCs, as shown by binding of wheat germ agglutinin and a PMC-specific monoclonal antibody. Cell transplantation and cell marking experiments have been carried out to determine the number of SMCs that convert when intermediate numbers of PMCs are present in the embryo. These experiments indicate that the number of converting SMCs is inversely proportional to the number of PMCs in the blastocoel. In addition, they show that PMCs and converted SMCs cooperate to produce a skeleton that is correct in both size and configuration. This regulatory system should shed light on the nature of cell-cell interactions that control cell differentiation and on the way in which evolutionary processes modify developmental programs.